1. Field of the Invention
This invention relates generally to mechanisms of gene expression in plants and more specifically to regulation of expression of genes in plants in a "tissue-preferred" manner. Regulation of expression is achieved using a transcriptional regulatory unit capable of driving expression of genes within certain tissues of a plant. Said transcriptional regulatory unit will ultimately be utilized for driving expression of genes that confer a selective advantage to a plant.
2. Description of the Related Art
Transcriptional control elements that drive "tissue-general" or "constitutive" gene expression in plants have been described. Examples include the promoters of the Agrobacterium nopaline synthase gene (Depicker, et al. 1982) and the maize ubiquitin gene (Christensen, et al. 1992). These promoters have been well characterized and utilized for driving gene expression in transgenic plants [e.g., CaMV 35S (Odell et al. 1985)]. There exists both an increasing interest in co-transforming plants with multiple plant transcription units and a realization of several potential problems associated with this technique. Concerns associated with the utilization of common regulatory sequences to drive expression of multiple genes include, but are not restricted to: a) recombination resulting from pairing along homologous regions, cross-overs and loss of the intervening region either prior to (in the case of a plasmid) or post-integration; b) hairpin loops caused by two copies of the sequence in opposite orientation adjacent to each other, again with possibilities of excision and loss of these regulatory regions; c) competition among different copies of the same promoter region for binding of promoter-specific transcription factors or other regulatory DNA-binding proteins; d) the relative strengths of expression of different promoters either within or between species, wherein one promoter may provide optimum levels of expression for one gene in a certain cell type or species, but may be either too strong or too weak for providing the required level of expression of a different gene in a certain cell type or species.
As part of our efforts to provide mechanisms for regulating the expression of genes that will ultimately be used for control of insect pests, primarily Ostrinia nubilalis, the European corn borer (ECB), we have been isolating and/or characterizing clones that exhibit intermediate to strong expression in the tissues that ECB primarily feeds on or tunnels through during its life cycle; these tissues include leaves, the leaf collar, the stalk rind and pith, and in the case of second generation ECB, pollen (Showers et al., 1989). While constitutive or "non-tissue-preferred" promoters have been demonstrated to be effective for this purpose such as the CAMV 35S promoter driving the Bacillus thuringiensis cryIA(b) gene provided effective ECB control in the field (Koziel et al., 1993), there are several advantages to utilizing promoters that function in a tissue-preferred manner. These include reduced resource drain on the plant in making a gene product constitutively, as well as localization or compartmentalization of gene expression in cases where the gene product must to be restricted to, or from, a certain tissue(s). Said gene products may include general cellular inhibitors such as RNases or other cytotoxins. As an example, Mariani, et al (Nature 347:737, 1990) engineered vectors that exhibited anther-specific gene expression of suc inhibitor genes for use in male sterility systems, since expression in regions other that the anther in a plant would be toxic. As an example of tissue-preferred expression, Koziel et al. (1993) utilized a combination of the maize PEP carboxylase promoter and a pollen promoter each driving cryIA (b) expression in separate constructs resulting in the generation of ECB-tolerant corn plants.
There is a need in the art for novel transcriptional regulatory elements which are capable of driving tissue-preferred gene expression in plants. It is considered important by those skilled in the art to continue to provide tissue-preferred transcription units capable of driving expression of genes that may confer a selective advantage to a plant.